Sugar cane (Saccharum spp.) is a highly polyploid plant grown in different parts of the world from the tropics to the subtropics, and accounts for around 60% of the world's sugar. It is also one of the important cash crops in many developing/developed countries, with a high trade value. The importance of sugar cane has increased in recent years because cane is an important raw material for sugar industries and allied industries producing alcohol, acetic acid, butanol, paper, plywood, industrial enzymes and animal feed. Considering its importance in the agricultural industry, concerted efforts are being made for its improvement using biotechnological approaches.
Agrobacterium tumefaciens is a soil-borne pathogen that is widely used to introduce heterologous polynucleotides into plant cells, including plant cells from sugar cane. A. tumefaciens transfers a particular polynucleotide segment of a tumor-inducing (Ti) plasmid into the nucleus of infected host cells, which subsequently stably integrates into the host's genome. Advantageously, heterologous polynucleotides can be placed between the borders of the Ti plasmid, or plasmid modified for this purpose, and transferred to plant cells.
Although Agrobacterium-mediated transformation has been used for genetic manipulation of sugar cane, efficiency and reproducibility of the available methodologies continue to be a challenge. In fact, A. tumefaciens induces necrosis in cultured, transformed sugar cane tissue, with a resultant low transformation frequency (Arencibia et al. (1998) Transgenic Res. 7:123-222; Enriquez-Obregón et al. (1997) Biotecnologia Aplicada 14:169-174; and de la Riva et al. (1998) Electron. J. Biotechno. 1:118-133).
Because of the importance of manipulating sugar cane for improved characteristics (e.g., increased resistance to biotic or abiotic stresses, or improved production), there is a need for additional methods that advantageously increase the efficiency of Agrobacterium-mediated transformation of this important agricultural crop.
Accordingly, the present inventors have overcome the deficiencies of the prior art by providing methods of Agrobacterium-mediated transformation of sugar cane that result in greater transformation efficiencies